The scope of the present invention includes compounds and pharmaceutical compositions useful as anti-inflammatory agents.
Studies have shown that endogenous electrophilic unsaturated-nitrated fatty acids, including nitro-linoleic acid (LNO2) and nitro-oleic acid (OA-NO2), can mediate anti-inflammatory and pro-survival signaling reactions [1]. The basis for the mediation are mainly produced because the presence of the nitro group on the double bond turns the β-carbon adjacent to the nitroalkene strongly electrophilic and reacts covalently with nucleophiles both in proteins (thiols and histidine residues) and low molecular weight molecules via Michael addition reactions [2].
Thus, biological electrophiles have emerged as mediators protecting against xenobiotic and oxidant injury. The transcription factor Nrf2 (nuclear factor erythroid 2-related factor 2)/Keap1 (Kelch-like ECH-associating protein) pathway mediates phase 2 gene activation [3]. Under normal conditions, Nrf2 localizes to the cytoplasmic suppressor protein Keap1 which has several critical cysteine residues that serve as sensors to environmental stresses such as ROS and electrophiles [3, 4]. Keap1 cysteines are oxidized or alkylated, causing a conformational change and liberating Nrf2 to translocate to the nucleus, bind to the cis-acting DNA regulatory antioxidant response element (ARE) and thereby transactivating Nrf2-dependent gene transcription [3, 4]. This includes enzymes involved in glutathione (GSH) metabolism, such as the subunits of the rate-limiting enzyme of glutathione synthesis, glutamate-cysteine ligase catalytic (GCLC) and modifier (GCLM) subunit genes. Also NAD(P)H:quinone oxidoreductase-1 (NQO1), which not only detoxifies xenobiotic quinones, but also reduces antioxidants vitamin E and coenzyme Q10 to their active form, is a Nrf2 target gene.
Additionally, HO-1 has been shown to be positively regulated by Nrf2 [5, 6]. This widespread mechanism protects against metabolic and inflammatory stress [5, 7, 8]. It is interesting to note that electrophilic nitro-fatty acids activate NRF2 by a KEAP1 cysteine 151-independent mechanism [9]. Actually, nitrated oleic acid, one of the endogenous nitroalkenes, is a Cys(151)-independent Nrf2 activator, which in turn can influence the pattern of gene expression and therapeutic actions of nitroalkenes [9].
Heme Oxygenase-1 (HO-1) also plays a central role in vascular inflammatory signaling and mediates a protective response to inflammatory stresses such as atherosclerosis, vascular restenosis and kidney diseases including transplant rejection [10]. Heme oxygenase 1 catalyzes the degradation of heme to biliverdin, iron, and carbon monoxide (CO). CO has been shown to display diverse, adaptive biological properties, including anti-inflammatory, anti-apoptotic, and vasodilatory actions [11].
Nitrated fatty acids have also been shown to be activators of peroxisome proliferator-activated receptor gamma (PPARγ). PPARγ is established as a master regulator of metabolism, inflammation, adipogenesis, and insulin sensitization [12]. High, non-physiological concentrations of native fatty acids (N50 μM), prostaglandin metabolites, and oxidized fatty acid derivatives are able to activate PPARγ, α, and δ [13,14]. Fatty acids containing an α-β-unsaturated ketone as a core structural element, such as 15d-PGJ2, also activate PPARγ [15]. Docking of 15d-PGJ2 to the ligand binding domain (LBD) shows that it is not sufficient to activate the receptor; rather, a covalent Michael addition reaction (locking reaction) is required for activation [13]. The PPARγ receptor contains a critical thiol (Cys285) in the LBD, with covalent modification of this highly conserved Cys285 by thiol-reactive compounds sufficient to induce partial receptor activation [16]. NO2-FAs and keto-fatty acid derivatives have high binding affinities for PPAR isotypes, being PPARγ the most robustly-activated receptor [13,17]. The mechanism by which LNO2 activates PPARγ has been determined with recent solution of the crystal structure of PPARγ having LNO2 occupancy in the LBD [18,19]. Differential conformational changes to PPARγ resulting from this unique endogenous ligand have the capacity to impart unique specificity to the downstream signaling events resulting from PPARγ activation [13,19]. Interestingly, PPARγ activation skews human monocytes toward an anti-inflammatory M2 phenotype [20], another possible mechanism to further explain the anti-inflammatory, anti-atherogenic properties of endogenous nitroalkenes.
However, in vivo studies of nitrated fatty acids such as nitrated oleic acid (18:1-NO2) metabolism showed that nitrated oleic acid undergoes a rapid and substantial modification that affects subsequent chemical reactivity and signaling actions [21]. More specifically, the results of the study showed the 18:1-NO2 suffers rapid but reversible adduction to plasma thiols and GSH. Furthermore, a significant proportion of 18:1-NO2 and its metabolites are converted to nitroalkane derivatives by saturation of the double bond, and to a lesser extent are desaturated to diene derivatives. The rapid saturation of the double bond decreases the electrophilic character of the molecule and may consequently affect the potency [22].
The study also showed that the hydrophobic nitro-oleic acid is metabolized by the β-oxidation pathways. As a result, the β-oxidized metabolite will be less hydrophobic and this will not only influence partitioning between hydrophobic and hydrophilic compartments and consequent tissue distribution, but can also affect chemical reactivity and pharmacological profiles by altering the specificity of the nitroalkenes to the biologically relevant targets.
Non-endogenous hydrophobic nitroalkene tocopherols and analogs thereof, as shown in WO 2015/073527, exhibit comparable potency as anti-inflammatory nitrated fatty acids and mimics transport processes of other lipid molecules in vivo and is closely related to lipoprotein homeostasis and metabolism which control intestinal absorption, traffic through the vascular compartment, and cellular uptake [20]. However, poor hydrosolubility of the nitroalkene tocopherol presents difficulties for traditional routes of drug delivery. Consequently, as discussed, in vivo distribution is premised on simulating the transport mechanism of endogenous lipid molecules.
A new family of nitroalkenes trolox (the hydrosoluble form of alpha-tocopherol) derivatives, provides advantages of controlling hydrosolubility, in doing so the scope of the invention includes hydrosoluble nitroalkene trolox derivatives to very hydrophobic ones. This allows the new generation of nitroalkenes to be modified for different inflammation related conditions.